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The aim of this investigation was to determine the effect of percutaneous neuromuscular stimulatory (PNS) pulse duration on whole muscle contractile properties of the human biceps brachii muscle as measured by a laser-based mechanomyographic (MMG) technique. This study quantified the effect of PNS pulse duration on muscle contractile properties to facilitate the development of standardized techniques to utilize MMG as an in-clinic assessment tool. Ten male volunteers, aged 19 to 33 years, participated in the study. Percutaneous neuromuscular stimulation (maximum of 180V and 85mA) was delivered by a transcutaneous electrical neuromuscular stimulation (TENS) device, through bipolar surface electrodes, to the short head of the right bicep brachii. The duration of the PNS pulse was randomly varied so that 10 trials were recorded at each of 10 PNS pulse durations, ranging from 50 micros, at 50-micros increments. In total, 100 trials for each subject were available for analysis. A laser MMG sensor detected the lateral displacement of the muscle belly during the development of isometric tension after each PNS pulse had been delivered. The parabolic MMG waveforms were analysed and statistically compared to determine whether PNS pulse duration influenced the recorded contractile properties of the biceps brachii. The results showed that the duration of the PNS pulse significantly (P<0.05) influenced the measured contractile properties of the biceps brachii as determined by the MMG technique. At PNS pulse durations below 300 micros, the contractile properties of the biceps brachii were more variable and more consistent with a slower contracting, or inadequately stimulated, muscle. At PNS pulse durations above 300 micros, the contractile properties of the muscle were considered stable and were similar to those previously reported for biceps brachii using a tensiometric technique. In conclusion, PNS pulse durations above 300 micros provide both a maximal lateral displacement of the muscle's belly and stable measures of its contractile properties. [ABSTRACT FROM AUTHOR]

Improving energy provision in the failing heart by augmenting the creatine kinase (CK) system is a desirable therapeutic target. However, over-expression of the creatine transporter (CrT-OE) has shown that very high creatine levels result in cardiac hypertrophy and dysfunction. We hypothesise this is due to insufficient endogenous CK activity to maintain thermodynamically favourable metabolite ratios. If correct, then double transgenic mice (dTg) overexpressing both CrT and the muscle isoform of CK (CKM-OE) would rescue the adverse phenotype. In Study 1, overexpressing lines were crossed and cardiac function assessed by invasive haemodynamics and echocardiography. This demonstrated that CKM-OE was safe, but too few hearts had creatine in the toxic range. In Study 2, a novel CrT-OE line was generated with higher, homogeneous, creatine levels and phenotyped as before. Myocardial creatine was 4-fold higher in CrT-OE and dTg hearts compared to wildtype and was associated with hypertrophy and contractile dysfunction. The inability of dTg hearts to rescue this phenotype was attributed to downregulation of CK activity, as occurs in the failing heart. Nevertheless, combining both studies in a linear regression analysis suggests a modest positive effect of CKM over a range of creatine concentrations. In conclusion, we confirm that moderate elevation of creatine is well tolerated, but very high levels are detrimental. Correlation analysis lends support to the theory that this may be a consequence of limited CK activity. Future studies should focus on preventing CKM downregulation to unlock the potential synergy of augmenting both creatine and CK in the heart., (Copyright © 2024. Published by Elsevier Ltd.)

Background: Phosphorus cardiovascular magnetic resonance spectroscopy ( 31 P-CMRS) has emerged as an important tool for the preclinical assessment of myocardial energetics in vivo. However, the high rate and diminutive size of the mouse heart is a challenge, resulting in low resolution and poor signal-to-noise. Here we describe a refined high-resolution 31 P-CMRS technique and apply it to a novel double transgenic mouse (dTg) with elevated myocardial creatine and creatine kinase (CK) activity. We hypothesised a synergistic effect to augment energetic status, evidenced by an increase in the ratio of phosphocreatine-to-adenosine-triphosphate (PCr/ATP)., Methods and Results: Single transgenic Creatine Transporter overexpressing (CrT-OE, n = 7) and dTg mice (CrT-OE and CK, n = 6) mice were anaesthetised with isoflurane to acquire 31 P-CMRS measurements of the left ventricle (LV) utilising a two-dimensional (2D), threefold under-sampled density-weighted chemical shift imaging (2D-CSI) sequence, which provided high-resolution data with nominal voxel size of 8.5 µl within 70 min. ( 1 H-) cine-CMR data for cardiac function assessment were obtained in the same imaging session. Under a separate examination, mice received invasive haemodynamic assessment, after which tissue was collected for biochemical analysis. Myocardial creatine levels were elevated in all mouse hearts, but only dTg exhibited significantly elevated CK activity, resulting in a 51% higher PCr/ATP ratio in heart (3.01 ± 0.96 vs. 2.04 ± 0.57-mean ± SD; dTg vs. CrT-OE), that was absent from adjacent skeletal muscle. No significant differences were observed for any parameters of LV structure and function, confirming that augmentation of CK activity does not have unforeseen consequences for the heart., Conclusions: We have developed an improved 31 P-CMRS methodology for the in vivo assessment of energetics in the murine heart which enabled high-resolution imaging within acceptable scan times. Mice over-expressing both creatine and CK in the heart exhibited a synergistic elevation in PCr/ATP that can now be tested for therapeutic potential in models of chronic heart failure., (© 2023. The Author(s).)

Aims: Low levels of homoarginine and creatine are associated with heart failure severity in humans, but it is unclear to what extent they contribute to pathophysiology. Both are synthesized via L-arginine:glycine amidinotransferase (AGAT), such that AGAT -/- mice have a combined creatine and homoarginine deficiency. We hypothesized that this would be detrimental in the setting of chronic heart failure., Methods and Results: Study 1: homoarginine deficiency-female AGAT -/- and wild-type mice were given creatine-supplemented diet so that both had normal myocardial creatine levels, but only AGAT -/- had low plasma homoarginine. Myocardial infarction (MI) was surgically induced and left ventricular (LV) structure and function assessed at 6-7 weeks by in vivo imaging and haemodynamics. Study 2: homoarginine and creatine-deficiency-as before, but AGAT -/- mice were given creatine-supplemented diet until 1 week post-MI, when 50% were changed to a creatine-free diet. Both groups therefore had low homoarginine levels, but one group also developed lower myocardial creatine levels. In both studies, all groups had LV remodelling and dysfunction commensurate with the development of chronic heart failure, for example, LV dilatation and mean ejection fraction <20%. However, neither homoarginine deficiency alone or in combination with creatine deficiency had a significant effect on mortality, LV remodelling, or on any indices of contractile and lusitropic function., Conclusions: Low levels of homoarginine and creatine do not worsen chronic heart failure arguing against a major causative role in disease progression. This suggests that it is unnecessary to correct hArg deficiency in patients with heart failure, although supra-physiological levels may still be beneficial., (© 2022 The Authors. ESC Heart Failure published by John Wiley & Sons Ltd on behalf of European Society of Cardiology.)

Organisms obtain creatine from their diet or by de novo synthesis via AGAT (L-arginine:glycine amidinotransferase) and GAMT (Guanidinoacetate N-methyltrasferase) in kidney and liver, respectively. AGAT also synthesizes homoarginine (hArg), low levels of which predict poor outcomes in human cardiovascular disease, while supplementation maintains contractility in murine heart failure. However, the expression pattern of AGAT has not been systematically studied in mouse tissues and nothing is known about potential feedback interactions between creatine and hArg. Herein, we show that C57BL/6J mice express AGAT and GAMT in kidney and liver respectively, whereas pancreas was the only organ to express appreciable levels of both enzymes, but no detectable transmembrane creatine transporter ( Slc6A8 ). In contrast, kidney, left ventricle (LV), skeletal muscle and brown adipose tissue must rely on creatine transporter for uptake, since biosynthetic enzymes are not expressed. The effects of creatine and hArg supplementation were then tested in wild-type and AGAT knockout mice. Homoarginine did not alter creatine accumulation in plasma, LV or kidney, whereas in pancreas from AGAT KO, the addition of hArg resulted in higher levels of tissue creatine than creatine-supplementation alone ( P < 0.05). AGAT protein expression in kidney was downregulated by creatine supplementation ( P < 0.05), consistent with previous reports of end-product repression. For the first time, we show that hArg supplementation causes a similar down-regulation of AGAT protein ( P < 0.05). These effects on AGAT were absent in the pancreas, suggesting organ specific mechanisms of regulation. These findings highlight the potential for interactions between creatine and hArg that may have implications for the use of dietary supplements and other therapeutic interventions., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Lygate, Lake, McAndrew, Neubauer and Zervou.)

Submaximal eccentric (ECC) cycling exercise is commonly used in research studies. No previous study has specified the required time naïve participants take to familiarize with submaximal ECC cycling. Therefore, we designed this study to determine whether critical indicators of cycling reliability and variability stabilize during 15 min of submaximal, semi-recumbent ECC cycling (ECC cycling). Twenty-two participants, aged between 18-51 years, volunteered to complete a single experimental session. Each participant completed three peak eccentric torque protocol (PETP) tests, nine countermovement jumps and 15 min of submaximal (i.e., 10% peak power output produced during the PETP tests) ECC cycling. Muscle activation patterns were recorded from six muscles (rectus femoris, RF; vastus lateralis, VL; vastus medialis, VM; soleus, SOL; medial gastrocnemius, GM; tibialis anterior, TA), during prescribed-intensity ECC cycling, using electromyography (EMG). Minute-to-minute changes in the reliability and variability of EMG patterns were examined using intra-class correlation coefficient (ICC) and variance ratios (VR). Differences between target and actual power output were also used as an indicator of familiarization. Activation patterns for 4/6 muscles (RF, VL, VM and GM) became more consistent over the session, the RF, VL and VM increasing from moderate (ICC = 0.5-0.75) to good (ICC = 0.75-0.9) reliability by the 11th minute of cycling and the GM good reliability from the 1st minute (ICC = 0.79, ICC range = 0.70-0.88). Low variability (VR ≤ 0.40) was maintained for VL, VM and GM from the 8th, 8th and 1st minutes, respectively. We also observed a significant decrease in the difference between actual and target power output (χ 2 14 = 30.895, p = 0.006, W = 0.105), expressed primarily between the 2nd and 3rd minute of cycling ( Z = -2.677, p = 0.007). Indicators of familiarization during ECC cycling, including deviations from target power output levels and the reliability and variability of muscle activation patterns stabilized within 15 min of cycling. Based upon this data, it would be reasonable for future studies to allocate ∼ 15 min to familiarize naïve participants with a submaximal ECC cycling protocol., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Walsh, McAndrew, Shemmell and Stapley.)

Eccentric (ECC) cycling is used in rehabilitation and sports conditioning settings. We present the construction and mode of operation of a custom-built semi-recumbent ECC cycle designed to limit the production of lower limb muscle activity to the phase of the pedal cycle known to produce ECC contractions. A commercially available semi-recumbent frame and seat (Monarch, 837E Semi-recumbent Bike, Sweden) were used to assemble the ergometer. An electrical drive train system was constructed using individual direct drive servo motors. To avoid active muscle activation occurring during the non-ECC pedaling phase of cycling, a "trip" mechanism was integrated into the drivetrain system using a servo-driven regenerative braking mechanism based on the monitoring of the voltage produced over and above a predetermined threshold produced by the motors. The servo drive internal (DC bus) voltage is recorded and internally monitored during opposing (OPP) and non-opposing (N-OPP) phases of the pedal cycle. To demonstrate that the cycle functions as desired and stops or "trips" when it is supposed to, we present average (of 5 trials) muscle activation patterns of the principal lower limb muscles for regular ECC pedal cycles in comparison with one pedal cycle during which the muscles activated outside the desired phase of the cycle for a sample participant. This semi-recumbent ECC cycle ergometer has the capacity to limit the occurrence of muscle contraction only to the ECC phase of cycling. It can be used to target that mode of muscle contraction more precisely in rehabilitation or training studies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Walsh, McAndrew, Henness, Shemmell, Cuicuri and Stapley.)

Information about the use and donor site morbidity of periosteal free flaps in head and neck reconstruction is limited. The aim of this study was to examine potential periosteal free flap donor sites with respect to their dimensions, tissue and pedicle characteristics, and predicted donor site morbidity in a cadaveric model. The following cadaveric periosteal specimens with a vascular pedicle were harvested using standard surgical approaches: skull, chest wall, sternum, scapula, iliac crest, femur, and humerus. Data relating to the periosteum size and quality, vascular pedicle, surgical factors, feasibility of use, and the potential donor-site morbidity were recorded. One female (age: 78 years, height: 152 cm) and one male (age: 65 years, height: 186 cm) cadaver were used for flap harvest. The skull, chest wall, scapula, and femur were suitable in terms of the size of the periosteum harvested. The procedure to remove the periosteum from the scalp, chest wall, and scapula had the least predicted donor-site morbidity. The pedicle length and vessel caliber from the periosteal flaps were most favorable from the skull, scapula, and iliac crest. Considering all factors, the periosteum harvested from the skull and scapula were the most promising., (Copyright © 2021 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of The American Society of Plastic Surgeons.)

Semi-recumbent eccentric (ECC) cycling is increasingly used in studies of exercise with healthy and clinical populations. However, workloads are generally prescribed using measures obtained during regular concentric cycling. Therefore, the purpose of the study was to quantify the reliability of measures derived from a protocol that elicited peak ECC torque produced by the lower limb in a semi-recumbent position. Experiments were carried out on a dynamometer in a seated, semi-recumbent position identical to that of a custom-built ECC cycle, a modified Monark recumbent cycle. Thirty healthy participants completed two testing sessions. Each session comprised three series of six repetitions of a peak ECC torque protocol (PETP) on an isokinetic dynamometer. Absolute and relative reliability of peak torque, power, angle of peak torque, and work (recorded for each repetition) was determined using coefficient of variation (CV) and intraclass correlation coefficient (ICC), respectively. Ratings of perceived exertion (RPE), muscle soreness, and perceived effort (PE) were recorded pre-PETP, immediately post-PETP, and 1-min post each PETP. The protocol showed absolute reliability values <15% for mean peak (CV = 10.6-12.1) torque, power (CV = 10.4-12.3), angle of peak torque (CV = 1.2-1.4), and work (CV = 9.7-12.1). Moderate to high between-test relative reliability is reported for mean and highest torque (ICC = 0.84-0.95; ICC = 0.88-0.98), power (ICC = 0.84-0.94; ICC = 0.89-0.98), and work (ICC = 0.84-0.93; ICC = 0.88-0.98), respectively. Within-session peak torque, peak power, and peak work showed high relative reliability for mean (ICC = 0.92-0.95) and highest (ICC = 0.92-0.97) values. Overall, the PETP test provides a reliable way of determining peak ECC torque specific to semi-recumbent ECC cycling that may be used to prescribe workloads for this form of exercise., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Walsh, Stapley, Shemmell and McAndrew.)

Aims: Adenylate kinase 1 (AK1) catalyses the reaction 2ADP ↔ ATP + AMP, extracting extra energy under metabolic stress and promoting energetic homeostasis. We hypothesised that increased AK1 activity would have negligible effects at rest, but protect against ischaemia/reperfusion (I/R) injury., Methods and Results: Cardiac-specific AK1 overexpressing mice (AK1-OE) had 31% higher AK1 activity ( P = 0.009), with unchanged total creatine kinase and citrate synthase activities. Male AK1-OE exhibited mild in vivo dysfunction at baseline with lower LV pressure, impaired relaxation, and contractile reserve. LV weight was 19% higher in AK1-OE males due to higher tissue water content in the absence of hypertrophy or fibrosis. AK1-OE hearts had significantly raised creatine, unaltered total adenine nucleotides, and 20% higher AMP levels ( P = 0.05), but AMP-activated protein kinase was not activated ( P = 0.85). 1 H-NMR revealed significant differences in LV metabolite levels compared to wild-type, with aspartate, tyrosine, sphingomyelin, cholesterol all elevated, whereas taurine and triglycerides were significantly lower. Ex vivo global no-flow I/R, caused four-of-seven AK1-OE hearts to develop terminal arrhythmia (cf. zero WT), yet surviving AK1-OE hearts had improved functional recovery. However, AK1-OE did not influence infarct size in vivo and arrhythmias were only observed ex vivo , probably as an artefact of adenine nucleotide loss during cannulation., Conclusion: Modest elevation of AK1 may improve functional recovery following I/R, but has unexpected impact on LV weight, function and metabolite levels under basal resting conditions, suggesting a more nuanced role for AK1 underpinning myocardial energy homeostasis and not just as a response to stress., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zervou, McAndrew, Whittington, Lake, Park, Cha, Ostrowski, Eykyn, Schneider, Neubauer and Lygate.)

Background: Advances in the control of transcranial magnetic stimulation (TMS) have enabled greater randomisation of stimulus intensity. It is unclear if such randomisation improves assessments of corticospinal excitability., New Method: We recorded the amplitude of TMS-induced motor evoked potentials (MEPs) from the first dorsal interosseous muscle of eleven participants, during three TMS protocols: blocks of increasing intensity (IB), randomised blocks (RB) and inter-stimulus randomisation (IR). Stimulus intensities from 90 to 140% of active motor threshold described corticospinal input-output (I/O) properties. The experiment was repeated in five participants., Results: Although MEP amplitudes did not differ between IB, RB and IR stimulation protocols, variability was lowest in the IR protocol, compared to IB and RB protocols. Reliability was highest in the IR protocol, compared to IB and IR protocols., Comparison With Existing Methods: Randomising TMS intensity between each trial produces less variable and more reliable estimates of corticospinal excitability than previously used blocked protocols and produces the same I/O measures., Conclusions: Inter-trial randomization of TMS intensities appears to be the most reliable method for constructing I/O curves at multiple time points and decreases the variability of responses., Competing Interests: Declaration of Competing Interest None., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Aim: Guanidinoacetate N -methyltransferase (GAMT) is the second essential enzyme in creatine (Cr) biosynthesis. Short-term Cr deficiency is metabolically well tolerated as GAMT -/- mice exhibit normal exercise capacity and response to ischemic heart failure. However, we hypothesized long-term consequences of Cr deficiency and/or accumulation of the Cr precursor guanidinoacetate (GA)., Methods: Cardiac function and metabolic profile were studied in GAMT -/- mice >1 year., Results: In vivo LV catheterization revealed lower heart rate and developed pressure in aging GAMT -/- but normal lung weight and survival versus age-matched controls. Electron microscopy indicated reduced mitochondrial volume density in GAMT -/- hearts ( P < 0.001), corroborated by lower mtDNA copy number ( P < 0.004), and citrate synthase activity ( P < 0.05), however, without impaired mitochondrial respiration. Furthermore, myocardial energy stores and key ATP homeostatic enzymes were barely altered, while pathology was unrelated to oxidative stress since superoxide production and protein carbonylation were unaffected. Gene expression of PGC-1α was 2.5-fold higher in GAMT -/- hearts while downstream genes were not activated, implicating a dysfunction in mitochondrial biogenesis signaling. This was normalized by 10 days of dietary Cr supplementation, as were all in vivo functional parameters, however, it was not possible to differentiate whether relief from Cr deficiency or GA toxicity was causative., Conclusion: Long-term Cr deficiency in GAMT -/- mice reduces mitochondrial volume without affecting respiratory function, most likely due to impaired biogenesis. This is associated with hemodynamic changes without evidence of heart failure, which may represent an acceptable functional compromise in return for reduced energy demand in aging mice., (Copyright © 2020 Aksentijević, Zervou, Eykyn, McAndrew, Wallis, Schneider, Neubauer and Lygate.)

Mitochondrial creatine kinase (Mt-CK) is a major determinant of cardiac energetic status and is down-regulated in chronic heart failure, which may contribute to disease progression. We hypothesised that cardiomyocyte-specific overexpression of Mt-CK would mitigate against these changes and thereby preserve cardiac function. Male Mt-CK overexpressing mice (OE) and WT littermates were subjected to transverse aortic constriction (TAC) or sham surgery and assessed by echocardiography at 0, 3 and 6 weeks alongside a final LV haemodynamic assessment. Regardless of genotype, TAC mice developed progressive LV hypertrophy, dilatation and contractile dysfunction commensurate with pressure overload-induced chronic heart failure. There was a trend for improved survival in OE-TAC mice (90% vs 73%, P = 0.08), however, OE-TAC mice exhibited greater LV dilatation compared to WT and no functional parameters were significantly different under baseline conditions or during dobutamine stress test. CK activity was 37% higher in OE-sham versus WT-sham hearts and reduced in both TAC groups, but was maintained above normal values in the OE-TAC hearts. A separate cohort of mice received in vivo cardiac 31 P-MRS to measure high-energy phosphates. There was no difference in the ratio of phosphocreatine-to-ATP in the sham mice, however, PCr/ATP was reduced in WT-TAC but preserved in OE-TAC (1.04 ± 0.10 vs 2.04 ± 0.22; P = 0.007). In conclusion, overexpression of Mt-CK activity prevented the changes in cardiac energetics that are considered hallmarks of a failing heart. This had a positive effect on early survival but was not associated with improved LV remodelling or function during the development of chronic heart failure.

Maintenance of upright standing posture has often been explained using the inverted pendulum model. This model considers the ankle plantarflexors to act as a single synergistic group. There are differences in muscle properties among the medial and lateral gastrocnemius (MG and LG, respectively) and the soleus that may affect their activation. Twelve volunteers participated in an investigation to determine whether the activation of the ankle plantarflexor muscles was modulated according to perturbation direction during unilateral standing perturbations of 1% body mass. High-density surface electromyography (HDS-EMG) was used to determine the amplitude and barycenter of the muscle activation and kinematic analysis was used to evaluate ankle, knee, and hip joint movement. The HDS-EMG amplitude and barycenter of MG and LG were modulated with the perturbation direction (MG p < 0.05; LG p < 0.01; one-way repeated-measures ANOVA). In soleus, the HDS-EMG barycenter modulated across the perturbation direction (p < 0.01 for X&Y coordinates), but the HDS-EMG amplitude did not change. A repeated-measures correlation was used to interpret the HDS-EMG pattern in the context of the kinematics. The relative contribution of MG activation compared to the total gastrocnemii activation was significantly associated with ankle dorsi/plantarflexion (r rm  = 0.620), knee flexion/extension and abduction/adduction (r rm  = 0.622 and r rm  = 0.547, respectively), and hip flexion/extension and abduction/adduction (r rm  = 0.653 and r rm  = 0.432, respectively). The findings suggest that the central nervous system activates motor units within different regions of MG, LG and SOL in response to standing perturbations in different directions.

This study investigated the effects of eccentric (ECC) and concentric (CON) semi-recumbent leg cycling on global corticospinal excitability (CSE), assessed through the activity of a non-exercised hand muscle. Thirteen healthy male adults completed two 30-min bouts of moderate intensity ECC and CON recumbent cycling on separate days. Power output (POutput), heart rate (HR) and cadence were monitored during cycling. Global CSE was assessed using transcranial magnetic stimulation to elicit motor-evoked potentials (MEP) in the right first dorsal interosseous muscle before ('Pre'), interleaved (at 10 and 20 mins, t10 and t20, respectively), immediately after (post, P0), and 30-min post exercise (P30). Participants briefly stopped pedalling (no more than 60 s) while stimulation was applied at the t10 and t20 time-points of cycling. Mean POutput, and rate of perceived exertion (RPE) did not differ between ECC and CON cycling and HR was significantly lower during ECC cycling (P = 0.01). Group mean MEP amplitudes were not significantly different between ECC and CON cycling at P0, t10, t20, and P30 and CON (at P > 0.05). Individual participant ratios of POutput and MEP amplitude showed large variability across the two modes of cycling, as did changes in slope of stimulus-response curves. These results suggest that compared to 'Pre' values, group mean CSE is not significantly affected by low-moderate intensity leg cycling in both modes. However, POutput and CSE show wide inter-participant variability which has implications for individual neural responses to CON and ECC cycling and rates of adaptation to a novel (ECC) mode. The study of CSE should therefore be analysed for each participant individually in relation to relevant physiological variables and account for familiarisation to semi-recumbent ECC leg cycling.

A capsule endoscopy examination of the human small bowel generates a large number of images that have high similarity. In order to reduce the time it takes to review the high similarity images, clinicians will increase the playback speed, typically to 15 frames per second [1]. Associated with this behaviour is an increased probability of overlooking an image that may contain an abnormality. An alternative option to increasing the playback speed is the application of abnormality detection systems to detect abnormalities such as ulcers, tumors, polyps and bleeding. However, applying all of these detection systems requires significant computing time and still produces numerous images with high similarity depending on the specificity of the utilized detection systems. An interesting approach to reduce viewing time is the application of a frame reduction system that reduces the number of images by omitting those with a high similarity of information. The advantage of such a system is that the specialist only needs to review a single image that technically represents a series of images with high similarity. This reduces the total number of images that a specialist must review and importantly, images containing any abnormality are not removed from the review, but simply reduced in number. Thus, the current study developed a frame reduction system using various color models using Bayer images for color texture and a modified local binary pattern (LBP) for structural information. The proposed system achieved a reduction ratio of 93.87%, which is higher than the existing systems and required lesser computation due to the utilization of Bayer images., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Aims: Mitochondrial creatine kinase (MtCK) couples ATP production via oxidative phosphorylation to phosphocreatine in the cytosol, which acts as a mobile energy store available for regeneration of ATP at times of high demand. We hypothesized that elevating MtCK would be beneficial in ischaemia-reperfusion (I/R) injury., Methods and Results: Mice were created over-expressing the sarcomeric MtCK gene with αMHC promoter at the Rosa26 locus (MtCK-OE) and compared with wild-type (WT) littermates. MtCK activity was 27% higher than WT, with no change in other CK isoenzymes or creatine levels. Electron microscopy confirmed normal mitochondrial cell density and mitochondrial localization of transgenic protein. Respiration in isolated mitochondria was unaltered and metabolomic analysis by 1 H-NMR suggests that cellular metabolism was not grossly affected by transgene expression. There were no significant differences in cardiac structure or function under baseline conditions by cine-MRI or LV haemodynamics. In Langendorff-perfused hearts subjected to 20 min ischaemia and 30 min reperfusion, MtCK-OE exhibited less ischaemic contracture, and improved functional recovery (Rate pressure product 58% above WT; P < 0.001). These hearts had reduced myocardial infarct size, which was confirmed in vivo: 55 ± 4% in WT vs. 29 ± 4% in MtCK-OE; P < 0.0001). Isolated cardiomyocytes from MtCK-OE hearts exhibited delayed opening of the mitochondrial permeability transition pore (mPTP) compared to WT, which was confirmed by reduced mitochondrial swelling in response to calcium. There was no detectable change in the structural integrity of the mitochondrial membrane., Conclusions: Modest elevation of MtCK activity in the heart does not adversely affect cellular metabolism, mitochondrial or in vivo cardiac function, but modifies mPTP opening to protect against I/R injury and improve functional recovery. Our findings support MtCK as a prime therapeutic target in myocardial ischaemia.

Aims: Creatine buffers cellular adenosine triphosphate (ATP) via the creatine kinase reaction. Creatine levels are reduced in heart failure, but their contribution to pathophysiology is unclear. Arginine:glycine amidinotransferase (AGAT) in the kidney catalyses both the first step in creatine biosynthesis as well as homoarginine (HA) synthesis. AGAT-/- mice fed a creatine-free diet have a whole body creatine-deficiency. We hypothesized that AGAT-/- mice would develop cardiac dysfunction and rescue by dietary creatine would imply causality., Methods and Results: Withdrawal of dietary creatine in AGAT-/- mice provided an estimate of myocardial creatine efflux of ∼2.7%/day; however, in vivo cardiac function was maintained despite low levels of myocardial creatine. Using AGAT-/- mice naïve to dietary creatine we confirmed absence of phosphocreatine in the heart, but crucially, ATP levels were unchanged. Potential compensatory adaptations were absent, AMPK was not activated and respiration in isolated mitochondria was normal. AGAT-/- mice had rescuable changes in body water and organ weights suggesting a role for creatine as a compatible osmolyte. Creatine-naïve AGAT-/- mice had haemodynamic impairment with low LV systolic pressure and reduced inotropy, lusitropy, and contractile reserve. Creatine supplementation only corrected systolic pressure despite normalization of myocardial creatine. AGAT-/- mice had low plasma HA and supplementation completely rescued all other haemodynamic parameters. Contractile dysfunction in AGAT-/- was confirmed in Langendorff perfused hearts and in creatine-replete isolated cardiomyocytes, indicating that HA is necessary for normal cardiac function., Conclusions: Our findings argue against low myocardial creatine per se as a major contributor to cardiac dysfunction. Conversely, we show that HA deficiency can impair cardiac function, which may explain why low HA is an independent risk factor for multiple cardiovascular diseases., (© The Author 2017 Published by Oxford University Press on behalf of the European Society of Cardiology.)

Sarcomere assembly is a highly orchestrated and dynamic process which adapts, during perinatal development, to accommodate growth of the heart. Sarcomeric components, including titin, undergo an isoform transition to adjust ventricular filling. Many sarcomeric genes have been implicated in congenital cardiomyopathies, such that understanding developmental sarcomere transitions will inform the aetiology and treatment. We sought to determine whether Thymosin β4 (Tβ4), a peptide that regulates the availability of actin monomers for polymerization in non-muscle cells, plays a role in sarcomere assembly during cardiac morphogenesis and influences adult cardiac function. In Tβ4 null mice, immunofluorescence-based sarcomere analyses revealed shortened thin filament, sarcomere and titin spring length in cardiomyocytes, associated with precocious up-regulation of the short titin isoforms during the postnatal splicing transition. By magnetic resonance imaging, this manifested as diminished stroke volume and limited contractile reserve in adult mice. Extrapolating to an in vitro cardiomyocyte model, the altered postnatal splicing was corrected with addition of synthetic Tβ4, whereby normal sarcomere length was restored. Our data suggest that Tβ4 is required for setting correct sarcomere length and for appropriate splicing of titin, not only in the heart but also in skeletal muscle. Distinguishing between thin filament extension and titin splicing as the primary defect is challenging, as these events are intimately linked. The regulation of titin splicing is a previously unrecognised role of Tβ4 and gives preliminary insight into a mechanism by which titin isoforms may be manipulated to correct cardiac dysfunction., (Copyright © 2016 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Mice over-expressing the creatine transporter have elevated myocardial creatine levels [Cr] and are protected against ischaemia/reperfusion injury via improved energy reserve. However, mice with very high [Cr] develop cardiac hypertrophy and dysfunction. To investigate these contrasting effects, we applied a non-biased hypothesis-generating approach to quantify global protein and metabolite changes in the LV of mice stratified for [Cr] levels: wildtype, moderately elevated, and high [Cr] (65-85; 100-135; 160-250 nmol/mg protein, respectively). Male mice received an echocardiogram at 7 weeks of age with tissue harvested at 8 weeks. RV was used for [Cr] quantification by HPLC to select LV tissue for subsequent analysis. Two-dimensional difference in-gel electrophoresis identified differentially expressed proteins, which were manually picked and trypsin digested for nano-LC-MS/MS. Principal component analysis (PCA) showed efficient group separation (ANOVA P ≤ 0.05) and peptide sequences were identified by mouse database (UniProt 201203) using Mascot. A total of 27 unique proteins were found to be differentially expressed between normal and high [Cr], with proteins showing [Cr]-dependent differential expression, chosen for confirmation, e.g. α-crystallin B, a heat shock protein implicated in cardio-protection and myozenin-2, which could contribute to the hypertrophic phenotype. Nuclear magnetic resonance (¹H-NMR at 700 MHz) identified multiple strong correlations between [Cr] and key cardiac metabolites. For example, positive correlations with α-glucose (r² = 0.45; P = 0.002), acetyl-carnitine (r² = 0.50; P = 0.001), glutamine (r² = 0.59; P = 0.0002); and negative correlations with taurine (r² = 0.74; P < 0.0001), fumarate (r² = 0.45; P = 0.003), aspartate (r² = 0.59; P = 0.0002), alanine (r² = 0.66; P < 0.0001) and phosphocholine (r² = 0.60; P = 0.0002). These findings suggest wide-ranging and hitherto unexpected adaptations in substrate utilisation and energy metabolism with a general pattern of impaired energy generating pathways in mice with very high creatine levels.

Background: The assessment of visual acuity is indicated in a number of clinical circumstances. It is commonly conducted through the use of a Snellen wall chart. Mobile technology developments and adoption rates by clinicians may potentially provide more convenient methods of assessing visual acuity. Limited data exist on the validity of these devices and applications., Objective: The objective of this study was to evaluate the assessment of distance visual acuity using mobile technology devices against the commonly used 3-metre Snellen chart in a primary care setting., Methods: A prospective quantitative comparative study was conducted at a regional medical practice. The visual acuity of 60 participants was assessed on a Snellen wall chart and two mobile technology devices (iPhone, iPad)., Results: Visual acuity intervals were converted to logarithm of minimum angle of resolution (logMAR) scores and subjected to intraclass correlation coefficient (ICC) assessment. The results show a high level of general agreement between testing modality (ICC 0.917 with a 95% confidence interval of 0.887-0.940)., Discussion: The high level of agreement of visual acuity results between the Snellen wall chart and both mobile technology devices suggests that clinicians can use this technology with confidence in the primary care setting.

Aims: Ischaemic heart disease is most prevalent in the ageing population and often exists with other comorbidities; however the majority of laboratory research uses young, healthy animal models. Several recent workshops and focus meetings have highlighted the importance of using clinically relevant models to help aid translation to realistic patient populations. We have previously shown that mice over-expressing the creatine transporter (CrT-OE) have elevated intracellular creatine levels and are protected against ischaemia-reperfusion injury. Here we test whether elevating intracellular creatine levels retains a cardioprotective effect in the presence of common comorbidities and whether it is additive to protection afforded by hypothermic cardioplegia., Methods and Results: CrT-OE mice and wild-type controls were subjected to transverse aortic constriction for two weeks to induce compensated left ventricular hypertrophy (LVH). Hearts were retrogradely perfused in Langendorff mode for 15 minutes, followed by 20 minutes ischaemia and 30 minutes reperfusion. CrT-OE hearts exhibited significantly improved functional recovery (Rate pressure product) during reperfusion compared to WT littermates (76% of baseline vs. 59%, respectively, P = 0.02). Aged CrT-OE mouse hearts (78±5 weeks) also had enhanced recovery following 15 minutes ischaemia (104% of baseline vs. 67%, P = 0.0007). The cardioprotective effect of hypothermic high K+ cardioplegic arrest, as used during cardiac surgery and donor heart transplant, was further enhanced in prolonged ischaemia (90 minutes) in CrT-OE Langendorff perfused mouse hearts (76% of baseline vs. 55% of baseline as seen in WT hearts, P = 0.02)., Conclusions: These observations in clinically relevant models further support the development of modulators of intracellular creatine content as a translatable strategy for cardiac protection against ischaemia-reperfusion injury.

New Findings: What is the central question of this study? There is an ethical imperative to optimize analgesia protocols for laboratory animals, but this is impeded by our inability to recognize pain reliably. We examined whether the Mouse Grimace Scale (MGS) provides benefits over a standard welfare scoring system for identifying a low level of pain in the frequently used murine surgical model of myocardial infarction. What is the main finding and its importance? Low-level pain, responsive to analgesia, was detected by MGS but not standard methods. In this model, most of the pain is attributable to the thoracotomy, excepted in mice with very large infarcts. This approach represents a model for assessing postsurgical analgesia in rodents. The Mouse Grimace Scale (MGS) was developed for assessing pain severity, but the general applicability to complex postsurgical pain has not been established. We sought to determine whether the MGS provides benefits over and above a standard welfare scoring system for identifying pain in mice following experimental myocardial infarction. Female C57BL/6J mice (n = 60), anaesthetized with isoflurane, were subjected to thoracotomy with ligation of a coronary artery or sham procedure. A single s.c. dose of buprenorphine (1.1 mg kg(-1) ) was given at the time of surgery and pain assessed at 24 h by MGS and a procedure-specific welfare scoring system. In some animals, a second dose of 0.6 mg kg(-1) buprenorphine was given and pain assessment repeated after 30 min. The MGS was scored from multiple photographs by two independent blinded observers with good correlation (r = 0.98). Using the average MGS score of both observers, we identified a subset of mice with low scores that were not considered to be in pain by the welfare scoring system or by single observer MGS. These mice showed a significant improvement with additional analgesia, suggesting that this low-level pain is real. Pain attributable to the myocardial injury, as opposed to thoracotomy, persisted at 24 h only in mice with large infarcts >40%. In conclusion, the use of a multi-observer, post hoc version of the MGS is a sensitive tool to assess the efficacy of postsurgical analgesic protocols. Following surgical induction of myocardial infarction, we identified a significant proportion of mice that were in low-level pain at 24 h that were not identified by other assessment methods., (© 2014 The Authors. Experimental Physiology published by John Wiley & Sons Ltd on behalf of The Physiological Society.)

Unlabelled: Inhibition of malonyl-coenzyme A decarboxylase (MCD) shifts metabolism from fatty acid towards glucose oxidation, which has therapeutic potential for obesity and myocardial ischemic injury. However, ~40% of patients with MCD deficiency are diagnosed with cardiomyopathy during infancy., Aim: To clarify the link between MCD deficiency and cardiac dysfunction in early life and to determine the contributing systemic and cardiac metabolic perturbations., Methods and Results: MCD knockout mice ((-/-)) exhibited non-Mendelian genotype ratios (31% fewer MCD(-/-)) with deaths clustered around weaning. Immediately prior to weaning (18days) MCD(-/-) mice had lower body weights, elevated body fat, hepatic steatosis and glycogen depletion compared to wild-type littermates. MCD(-/-) plasma was hyperketonemic, hyperlipidemic, had 60% lower lactate levels and markers of cellular damage were elevated. MCD(-/-) hearts exhibited hypertrophy, impaired ejection fraction and were energetically compromised (32% lower total adenine nucleotide pool). However differences between WT and MCD(-/-) converged with age, suggesting that, in surviving MCD(-/-) mice, early cardiac dysfunction resolves over time. These observations were corroborated by in silico modelling of cardiomyocyte metabolism, which indicated improvement of the MCD(-/-) metabolic phenotype and improved cardiac efficiency when switched from a high-fat diet (representative of suckling) to a standard post-weaning diet, independent of any developmental changes., Conclusions: MCD(-/-) mice consistently exhibited cardiac dysfunction and severe metabolic perturbations while on a high-fat, low carbohydrate diet of maternal milk and these gradually resolved post-weaning. This suggests that dysfunction is a common feature of MCD deficiency during early development, but that severity is dependent on composition of dietary substrates., (Copyright © 2014. Published by Elsevier Ltd.)

Background: Multiple studies suggest creatine mediates anti-oxidant activity in addition to its established role in cellular energy metabolism. The functional significance for the heart has yet to be established, but antioxidant activity could contribute to the cardioprotective effect of creatine in ischaemia/reperfusion injury., Objectives: To determine whether intracellular creatine levels influence responses to acute reactive oxygen species (ROS) exposure in the intact beating heart. We hypothesised that mice with elevated creatine due to over-expression of the creatine transporter (CrT-OE) would be relatively protected, while mice with creatine-deficiency (GAMT KO) would fare worse., Methods and Results: CrT-OE mice were pre-selected for creatine levels 20-100% above wild-type using in vivo (1)H-MRS. Hearts were perfused in isovolumic Langendorff mode and cardiac function monitored throughout. After 20 min equilibration, hearts were perfused with either H2O2 0.5 µM (30 min), or the anti-neoplastic drug doxorubicin 15 µM (100 min). Protein carbonylation, creatine kinase isoenzyme activities and phospho-PKCδ expression were quantified in perfused hearts as markers of oxidative damage and apoptotic signalling. Wild-type hearts responded to ROS challenge with a profound decline in contractile function that was ameliorated by co-administration of catalase or dexrazoxane as positive controls. In contrast, the functional deterioration in CrT-OE and GAMT KO hearts was indistinguishable from wild-type controls, as was the extent of oxidative damage and apoptosis. Exogenous creatine supplementation also failed to protect hearts from doxorubicin-induced dysfunction., Conclusions: Intracellular creatine levels do not influence the response to acute ROS challenge in the intact beating heart, arguing against creatine exerting (patho-)physiologically relevant anti-oxidant activity.

Changes in medical education have affected both curriculum design and delivery. Many medical schools now use integrated curricula and a systemic approach, with reduced hours of anatomy teaching. While learning anatomy via dissection is invaluable in educational, professional, and personal development, it is time intensive and supports a regional approach to learning anatomy; the use of prosections has replaced dissection as the main teaching method in many medical schools. In our graduate-entry medical degree, we use an integrated curriculum, with prosections to teach anatomy systemically. However, to not exclude dissection completely, and to expose students to its additional and unique benefits, we implemented a short "Dissection Experience" at the beginning of Year 2. Students attended three two-hour anatomy sessions and participated in dissection of the clinically relevant areas of the cubital fossa, femoral triangle, and infraclavicular region. This activity was voluntary and we retrospectively surveyed all students to ascertain factors influencing their decision of whether to participate in this activity, and to obtain feedback from those students who did participate. The main reasons students did not participate were previous dissection experience and time constraints. The reasons most strongly affecting students' decisions to participate related to experience (lack of previous or new) and new skill. Students' responses as to the most beneficial component of the dissection experience were based around practical skills, anatomical education, the learning process, and the body donors. We report here on the benefits and practicalities of including a short dissection experience in a systemic, prosection-based anatomy course., (© 2013 American Association of Anatomists.)

The aim of the present study was to determine how the intra-muscular segments of three shoulder muscles were coordinated to produce isometric force impulses around the shoulder joint and how muscle segment coordination was influenced by changes in movement direction, mechanical line of action and moment arm (ma). Twenty male subjects (mean age 22 years; range 18-30 years) with no known history of shoulder pathologies, volunteered to participate in this experiment. Utilising an electromyographic technique, the timing and intensity of contraction within 19 muscle segments of three superficial shoulder muscles (Pectoralis Major, Deltoid and Latissimus Dorsi) were studied and compared during the production of rapid (e.g. approximately 400ms time to peak) isometric force impulses in four different movement directions of the shoulder joint (flexion, extension, abduction and adduction). The results of this investigation have suggested that the timing and intensity of each muscle segment's activation was coordinated across muscles and influenced by the muscle segment's moment arm and its mechanical line of action in relation to the intended direction of shoulder movement (e.g. flexion, extension, abduction or adduction). There was also evidence that motor unit task groups were formed for individual motor tasks which comprise motor units from both adjacent and distant muscles. It was also confirmed that for any particular motor task, individual muscle segments can be functionally classified as prime mover, synergist or antagonist - classifications which are flexible from one movement to the next.